AFP Sensor Based on Ferrocene and AFPAb-Modified Electrode: Synthesis and Performance Evaluation

Two highly sensitive amperometric immunosensors for alpha-fetoprotein antigen (AFPAg) detection were fabricated through electrode modification strategies. Typically, during AFPAg detection, electron transfer is mediated by substances such as ferrocene or gold. However, in the case of the prepared zinc oxide electrodes, direct electron transfer was observed between the electrode surface and horseradish peroxidase (HRP). Specifically, by modifying a zinc electrode, pre-heated to 500°C, with AFP antibody (AFPAb) and ferrocene via chitosan, we developed two types of amperometric immunosensors for AFPAg quantification. Under optimized conditions, cyclic voltammetry revealed specific responses of the immunosensors to AFP within two distinct concentration ranges: 1.56 to 12.5 ng/mL and 25.0 to 100.0 ng/mL. The limits of detection were determined to be 0.02 ng/mL and 0.05ng/mL, respectively, at a signal-to-noise ratio of 3. The results demonstrated that electrodes modified with chitosan and AFPAb exhibited high sensitivity, indicating effective immobilization of AFPAb on the electrode surface and facilitating direct electron transfer between the electrode and HRP. Furthermore, when the electrode was additionally modified with ferrocene, a good sensitivity was achieved, suggesting that ferrocene also participated in electron transfer and contributed to signal amplification. We conducted a comprehensive investigation into the factors influencing the performance of the proposed immunosensors. The developed immunosensors exhibited excellent specificity, good reproducibility, repeatability, and long-term stability, making them promising candidates for clinical AFPAg detection.